The tooth root is critical for the function of our dentition because it anchors the tooth to the maxilla or mandible. In addition, the root helps transmit and balance occlusal forces through periodontal ligaments to the jaw bones and serves as a passageway for the neurovascular bundle that supplies blood flow, nutrition, and sensation to our teeth. Importantly, tooth root defects resulting from developmental malformations, pathological conditions and dental treatments are common and significantly compromise quality of life. However, currently we have limited information on the molecular and cellular regulatory mechanisms of crown-to-root transition and tooth root development. Significantly, we have recently identified epithelial and mesenchymal stem cells that are crucial for root development. BMP-mediated SHH signaling provides a niche for Sox2+ epithelial stem cells in molar root development. Moreover, our preliminary study indicated that BMP-mediated HH and FGF signaling in the epithelium may control the crown-to-root transition. We have identified Gli1+ cells as the dental mesenchymal stem cells responsible for controlling root development. In parallel, our preliminary results demonstrate that an epigenetic regulator Ezh2 modulates the transcriptional activity of key genes in the WNT and FGF pathways that are involved in regulating epithelial-mesenchymal interactions to control root formation. We have designed studies to test the hypothesis that BMP-mediated SHH, WNT and FGF signaling control the fate of epithelial and mesenchymal stem cells during root development. This signaling network and Ezh2 exert their functional specificity by regulating specific transcription factors and epithelial-mesenchymal interactions to control root patterning and development. Our Specific Aims are: (1) to examine how epithelial SHH and FGF, acting downstream of BMP signaling, control the fate of dental epithelial stem cells during root development; (2) to investigate the functional significance of WNT and FGF signaling in regulating specific transcription factors and the fate of CNC-derived MSCs during root development. We will elucidate the molecular mechanism(s) by which signals from the dental mesenchyme control root furcation and development; and (3) to investigate how Ezh2 controls the fate of CNC-derived MSCs during root development. We will investigate the Ezh2-mediated molecular signaling mechanism in regulating tissue-tissue interactions to control root patterning and development.